Dickkopf-1 and ROCK2 upregulation and associated protein aggregation in pseudoexfoliation syndrome and glaucoma

Abstract

AimsThe etiology of pseudoexfoliation (PEX), a stress-induced fibrillopathy and a leading cause of secondary glaucoma worldwide, remains limited. This study aims to understand the role of the Wnt antagonist Dickkopf-related protein 1 (DKK1) in PEX pathophysiology and assess its candidature as a biomarker for PEX. Main methodsExpression levels of DKK1 and Wnt signaling genes were assayed in the anterior ocular tissues of study subjects by qRT-PCR, Western blotting, and immunohistochemistry. Protein aggregation was studied through Proteostat staining. Role of DKK1 in protein aggregation and regulation of target Wnt signaling genes was elucidated through overexpression and knockdown studies in Human Lens Epithelial cells (HLEB3). Levels of DKK1 in circulating fluids were assayed through ELISA. Key findingsDKK1 upregulation was observed in lens capsule and conjunctiva tissues of PEX individuals compared to controls correlating with an upregulation of the Wnt signaling target, ROCK2. Proteostat staining showed increased protein aggregates in lens epithelial cells of PEX patients. HLE B-3 cells overexpressed with DKK1 showed increased protein aggregates along with upregulation of ROCK2, and knockdown of DKK1 in HLE B-3 cells demonstrated downregulation of ROCK2. Further, ROCK2 inhibition by Y-27632 in DKK1 overexpressed cells showed that DKK1 regulated protein aggregation via ROCK2. Also, increased levels of DKK1 were observed in patients' plasma and aqueous humor compared to controls. SignificanceThis study shows that DKK1 and ROCK2 might play a role in protein aggregation in PEX. Further, elevated levels of DKK1 in aqueous humor serve as a fair classifier of pseudoexfoliation glaucoma.